Summary

RMgm-4447
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_0718600; Gene model (P.falciparum): PF3D7_0416600; Gene product: conserved Plasmodium protein, unknown function (prohibitin-like protein, PHBL)
Transgene
Transgene not Plasmodium: mCherry
Promoter: Gene model: PBANKA_0711900; Gene model (P.falciparum): PF3D7_0818900; Gene product: heat shock protein 70 (HSP70)
3'UTR: Gene model: PBANKA_1340000; Gene product: dihydrofolate synthase/folylpolyglutamate synthase, putative (PbDHFS-FPGS)
Replacement locus: Gene model: PBANKA_0701500; Gene product: prohibitin, putative (PHB1)
Phenotype Asexual bloodstage; Fertilization and ookinete; Oocyst; Sporozoite;
Last modified: 27 April 2018, 18:24
  *RMgm-4447
Successful modificationThe parasite was generated by the genetic modification
The mutant contains the following genetic modification(s) Gene disruption, Introduction of a transgene
Reference (PubMed-PMID number) Reference 1 (PMID number) : 29669282
MR4 number
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone Not applicable
Other information parent line
The mutant parasite was generated by
Name PI/ResearcherMatz JM, Kooij TWA
Name Group/DepartmentDepartment of Molecular Parasitology, Institute of Biology
Name InstituteHumboldt University
CityBerlin
CountryGermany
Name of the mutant parasite
RMgm numberRMgm-4447
Principal namePBHL(-)
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Phenotype
Asexual blood stageReduced growth rate of asexual blood stages (the PHBL-deficient mutant grew at only 37% of the wild-type (WT) rate.
Reduced virulence (no cerebral complications in infected mice)
Gametocyte/GameteNot different from wild type
Fertilization and ookineteReduced formation of ookinetes in vitro. The ookinetes that were formed in vitro displayed normal gliding motility.
No ookinete formation could be detected in vivo (in midguts of A stephensi)
OocystNo oocyst formation
SporozoiteNo oocyst and sporozoite formation
Liver stageNot tested
Additional remarks phenotype

Mutant/mutation
The mutant lacks expression of PHBL and expresses mCherry under the control of the constitutive hsp70 promoter

Protein (function)
Proteins of the stomatin/prohibitin/flotillin/HfIK/C (SPFH) family are membrane-anchored and perform diverse cellular functions in different organelles. A systematic database search for Plasmodium SPFH proteins  uncovered four candidates that either display a close phylogenetic relationship to conserved members—prohibitin 1 (PHB1, PBANKA_0701500), prohibitin 2 (PHB2, PBANKA_1213100), and stomatin-like protein (STOML, PBANKA_0807100)- 0807100)—or fall into a unique sister group that contains related proteins from the phylum Myzozoa, which harbors the subphylum Apicomplexa. Accordingly,  the latter Plasmodium protein was named prohibitin-like protein (PHBL, PBANKA_0718600). All four proteins contain a central signature SPFH domain and a hydrophobic stretch Although the conventional Plasmodium SPFH proteins display a similarly high degree of conservation, PHBL sequences are rather divergent already among related Plasmodium species.

Phenotype
Reduced ookinete formation in vitro. No ookinetes could be detected in vivo (in midguts of A stephensi).

Additional information
Evidence is presented that:
- Mitochondrial Organization of PHBL-Deficient Parasites Remains Unaltered
- PHBL Regulates Mitochondrial Membrane Potential during Sexual Development


Other mutants


  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_0718600
Gene Model P. falciparum ortholog PF3D7_0416600
Gene productconserved Plasmodium protein, unknown function
Gene product: Alternative nameprohibitin-like protein, PHBL
Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/construct used(Linear) plasmid double cross-over
PlasmoGEM (Sanger) construct/vector usedNo
Modified PlasmoGEM construct/vector usedNo
Plasmid/construct map
Plasmid/construct sequence
Restriction sites to linearize plasmid
Partial or complete disruption of the geneComplete
Additional remarks partial/complete disruption
Selectable marker used to select the mutant parasitehdhfr/yfcu
Promoter of the selectable markerpbdhfr
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modification
Additional remarks selection procedure
Primer information: Primers used for amplification of the target sequences  Click to view information
Primer information: Primers used for amplification of the target sequences  Click to hide information
Sequence Primer 1
Additional information primer 1
Sequence Primer 2
Additional information primer 2
Sequence Primer 3
Additional information primer 3
Sequence Primer 4
Additional information primer 4
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6

  Transgene: Mutant parasite expressing a transgene
Type and details of transgene
Is the transgene Plasmodium derived Transgene: not Plasmodium
Transgene namemCherry
Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/construct(Linear) plasmid double cross-over
PlasmoGEM (Sanger) construct/vector usedNo
Modified PlasmoGEM construct/vector usedNo
Plasmid/construct map
Plasmid/construct sequence
Restriction sites to linearize plasmid
Selectable marker used to select the mutant parasitehdhfr/yfcu
Promoter of the selectable markerpbdhfr
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modification
Additional remarks selection procedure
Other details transgene
Promoter
Gene Model of Parasite PBANKA_0711900
Gene Model P. falciparum ortholog PF3D7_0818900
Gene productheat shock protein 70
Gene product: Alternative nameHSP70
Primer information details of the primers used for amplification of the promoter sequence  Click to view information
Primer information details of the primers used for amplification of the promoter sequence  Click to hide information
Sequence Primer 1
Additional information primer 1
Sequence Primer 2
Additional information primer 2
3'-UTR
Gene Model of Parasite PBANKA_1340000
Gene productdihydrofolate synthase/folylpolyglutamate synthase, putative
Gene product: Alternative namePbDHFS-FPGS
Primer information details of the primers used for amplification the 3'-UTR sequences  Click to view information
Primer information details of the primers used for amplification the 3'-UTR sequences  Click to hide information
Sequence Primer 1
Additional information primer 1
Sequence Primer 2
Additional information primer 2
Insertion/Replacement locus
Replacement / InsertionReplacement locus
Gene Model of Parasite PBANKA_0701500
Gene productprohibitin, putative
Gene product: Alternative namePHB1
Primer information details of the primers used for amplification of the target sequences  Click to view information
Primer information details of the primers used for amplification of the target sequences  Click to hide information
Sequence Primer 1
Additional information primer 1
Sequence Primer 2
Additional information primer 2
Sequence Primer 3
Additional information primer 3
Sequence Primer 4
Additional information primer 4